Obesity and overweight, which have been immense epidemics in developed countries for the last two decades, are associated with a wide array of diseases including cancer of different organs such as the colon. To date, the underlying mechanisms for these associations are not completely understood. One of emerging possibilities is that excess lipids accumulated in the tissue of obese individuals directly stimulates progression of colonic tumors. Intracellular lipids are stored in lipid droplets (LDs), dynamic organelles found in adipose tissue, as well as in many human tumors. It is believed that LDs are the source of energy that drives proliferation, a hallmark of tumor progression; however, their regulation and function are not well understood. Our preliminary data reveals that LD accumulation is increased in the colon of obese mice as well as in inflamed and neoplastic colonic tissue. Moreover, LDs are negatively regulated by tumor suppressor FOXO3, while stimulating LDs accumulation leads to a loss of FOXO3 activity. Thus we propose the existence of an LD-FOXO3 network that also includes PI3K and SIRT6 (a negative regulator of triglycerides (TGs). Moreover, preliminary data suggested that the LD-FOXO3 network is critical in proliferation of colonic cancer cells and also operates in obesity influence colonic epithelium. There is support that the LD-FOXO3 network is upstream control by EGFR and/or PGE2, while downstream modifies cell cycle regulators and triglyceride synthesis. The central hypothesis of this proposal is that proliferation in colonic cells controlled by LD-FOXO3 network leads to coordinate LD accumulation and cell cycle progression dependent on loss of FOXO3 is one mechanism responsible for obesity related colon tumor progression. The following Specific Aims are proposed: Aim 1. To assess if the LD induced proliferative response in colonic cancer cells depends on FOXO3 activity via (a) activation of EGFR and whether this is dependent on PGE2, (b) PI3K dependent loss of FOXO3 cell cycle arrest, and (c) SIRT6 dependent, a negative regulator of TGs in colon cancer cells. Aim 2. To determine if TNF-induced proliferation and consequent LD accumulation are due to activation of EGFR, PGE2 and a loss of FOXO3 cell cycle arrest and SIRT6 (in vitro), and (b) to find if selective blockade of LD in TNF treated Foxo3 or EGFR deficient mice suppresses proliferation of colonic epithelia by promoting Foxo3 dependent SIRT6 and cell cycle arrest and TGs. Aim 3. To assess if obesity and a deficiency in Foxo3 (a) exacerbated adenoma progression in DSS/AOM model leading to alterations in regulators of G0-G1 checkpoint and TGs, (b) if adenoma progression is suppressed by selective blockade of LD, PGE2, or TNF, and (c) to identify the Foxo3 dependent genes regulating lipid metabolism. Findings from the proposed study would have significant positive effects on human health by presenting the LD-FOXO3 network as a novel therapeutic target. Although mechanisms of colon cancer progression are complex, the inhibition of LDs may provide an effective blockade to tumor progression in patients with obesity promoted colonic tumor progression.